JMJD2C demethylates the transcriptionally repressive H3K9Me3 mark on the histone H3 within nucleosomes, which are the basic building blocks of chromatin. JMJD2C promotes cellular proliferation in several different types of cancer due to genomic amplification and overexpression. JMJD2C overexpression causes increased transcription of several oncogenes, including Mdm2, c-MYC, and NOTCH1. It is therefore possible that JMJD2C promotes oncgenesis by direct catalytic activity at the loci of target oncogenes. The objective of this work is to develop a potent and selective small molecule inhibitor of JMJD2C, and use this inhibitor to investigate the mechanism by which this demethylase impacts cancer cell proliferation and survival. We hypothesize that elaboration of a small-molecule fragment identified by molecular docking will enable the development of an effective JMJD2C inhibitor, and that inhibition of JMJD2C will exhibit selective toxicity to cancer cells by preventing the transcription of oncogenes regulated by this demethylase. In Aim 1 we will synthesize derivatives of our hit compound. In Aim 2 we will test the potency and selectivity of these inhibitors against a panel of demethylases and related proteins using in vitro inhibition assays in order to establish structure-activity relationship (SAR). Finally, in Aim 3 we will test he ability of selected compounds to inhibit the proliferation of cancer cells. We will also explore th mechanism of toxicity to cancer cells by determining changes in expression levels of cancer associated genes directly transcriptionally controlled by JMJD2C. The overall goal of this study is to develop a potent and selective small molecule inhibitor of JMJD2C that will be used in cells to validate JMJD2C as a therapeutic target for cancer, and to reveal mechanism(s) by which JMJD2C promotes oncogenesis.